Multiple enzyme cleaner for surgical instruments and endoscopes

ABSTRACT

The liquid composition is based on surfactants and enzymes, and is particularly useful for manual cleaning of instruments. In manual and ultrasonic application the composition in use concentration shows low foaming and gives a cleaning solution which is not cloudy at least at a temperature in the range from 16° C. to 40° C. In addition, the composition has good cleaning efficacy over a broad temperature range (20 to 55° C.) and shows good material compatibility.

CROSS REFERENCE TO RELATED APPLICATION

This application is a continuation of U.S. patent application Ser. No.12/642,091, filed Dec. 18, 2009, issued as U.S. Pat. No. 7,858,029,which is a continuation of U.S. patent application Ser. No. 12/351,027,filed Jan. 9, 2009, issued as U.S. Pat. No. 7,670,549, which is acontinuation of U.S. patent application Ser. No. 12/020,883, filed Jan.28, 2008, issued as U.S. Pat. No. 7,491,362, the entire disclosures ofwhich are incorporated herein by reference in their entirety.

The present invention relates generally to compositions and methods forcleaning surgical, medical and dental instruments prior to reuse, andmore particularly to treatment of the instruments prior to adisinfection process.

BACKGROUND OF THE INVENTION

Surgical, medical, and dental instruments after use are typicallycontaminated with blood and other body matter and potentially withinfectious microorganisms. Before being reused in a future procedurethese instruments must be washed and disinfected where indicated. Atypical cycle for cleaning medical instruments consists of a number ofconsecutive stages: pre-wash, wash, rinses (usually two) and drying. Thepre-wash stage is used to dissolve blood on the instruments and may berun with a wash solution containing detergent and possibly enzymes. Thewash part of the cycle is run with a wash solution containing detergentand possibly enzymes. Wash time, water temperature and detergentselection and concentration are matched according to requirements.Rinses are used to remove soil dissolved in the wash stage as well asthe remaining detergent.

The process of washing and disinfecting becomes complicated when bloodor other matter are allowed to dry on the instruments. The body fluids,such as blood, lipids and synovial fluids from joints adhere to theitems used during a procedure. As these fluids dry, the adhesion getsstronger and the fluids get harder to dissolve using ordinary cleaningmethods. Blood in particular becomes much more difficult to remove onceit has dried. Eventually, the adhesion of the soils becomes too strongfor normal detergents to break and the instruments remain soiled aftercleaning. The chemical structures of these detergents do not allow themto react with body fluids without the body fluids first being changed byother chemicals like enzymes. Enzymes like protease break these bodyfluids down by the chemical reaction called hydrolysis which also breaksdown their adhesive bond to the items the fluids are adhered to. Whenbroken down in this manner, body fluids become more soluble insurfactants and can then be washed away.

The compositions and methods currently used have the drawback that whenusing them in manual cleaning of instruments the personnel cannot seethe items to be cleaned and their contamination when these items areimmersed in the wash solution due to the foaming or cloudiness of thesolution. It is against this background that the present invention hasbeen made.

DETAILED DESCRIPTION OF THE INVENTION

The object of the present invention was to provide a detergentcomposition on the basis of surfactants and enzymes, which isparticularly useful for manual cleaning of instruments and endoscopes,which shows low foaming in use concentration, which gives a cleaning orwash solution which in use concentration is not cloudy at least at atemperature in the range from 16° C. to 40° C., and further has goodcleaning efficacy and material compatibility. This composition isadvantageous because it allows personnel to visually monitor thecleaning without foam or cloudy water interfering.

The present invention can be a liquid concentrate composition comprisingbased on the whole composition

-   a) 1 to 30 wt.-% of a low-foam surfactant system comprising at least    two different nonionic surfactants,    -   wherein one surfactant (i) is selected from a linear alkoxylated        fatty alcohol and an oxo alcohol of the formula R₁-A-OH,    -   wherein R₁ is a linear or branched C₁₀ to C₁₆ alkyl or alkenyl        group,    -   wherein A is —(OC₂H₄)_(x)—(OC₃H₆)_(y)— or        —(OC₃H₆)_(y)—(OC₂H₄)_(x)—, and    -   x is in the range from 7 to 9 (preferably 8), and    -   y is in the range from 1 to 3 (preferably 1 to 2), and has a        cloud point at a temperature in the range from 64 to 70° C.,    -   and the second surfactant (ii) is selected from a linear        alkoxylated fatty alcohol and an oxo alcohol of the formula        R₂-A-OH,    -   wherein R₂ is a linear or branched C₁₂ to C₁₅ alkyl or alkenyl        group,    -   wherein A is —(OC₂H₄)_(x)—(OC₃H₆)_(y)— or        —(OC₃H₆)_(y)—(OC₂H₄)_(x)—, and    -   x is in the range from 1 to 3 (preferably 2), and    -   y is in the range from 4 to 6 (preferably 5), and has a cloud        point at a temperature in the range from 39 to 41° C.;-   b) at least two different proteases, wherein preferably at least one    protease is a serine endopeptidase;-   c) lipase;-   d) optionally amylase;-   e) 10 to 70 wt.-% water and/or a compound of the general formula    X—CH₂—(CHY)_(n)—CH₂—Z (I), wherein n is an integer of from 0 to 2    and X, Y and Z independently symbolize hydrogen or a hydroxyl group    provided that at least 2 hydroxyl groups are present; and-   f) enzyme stabilizers.

The present invention is particularly useful for manual and ultrasoniccleaning of surgical instruments and endoscopes where visual inspectionis helpful. This liquid composition shows low foaming in useconcentration and gives a solution which in use concentration is notcloudy at a temperature in the range from 0 to 55° C., preferably from10 to 50° C. and further preferred from 16° C. to 40° C. Further, theliquid composition according to the present invention has good cleaningefficacy over a broad temperature range (20 to 55° C.) and shows goodmaterial compatibility. In addition the liquid detergent compositionaccording to the present invention also can be used for automatedapplication in washers at a temperature up to about 55° C.

The composition preferably has a pH in the range from 5.5 to 9.0, or inthe range from 6.0 to 8.5.

The present invention also provides a use dilution comprising theconcentrate according to the present invention diluted with water to apercentage ranging from about 85 to about 0.5 wt.-%.

Surfactant

The composition includes at least two surfactants.

One surfactant (i) is selected from a linear alkoxylated fatty alcoholand an oxo alcohol of the formula R₁-A-OH,

wherein R₁ is a linear or branched C₁₀ to C₁₆ alkyl or alkenyl group,

wherein A is —(OC₂H₄)_(x)—(OC₃H₆)_(y)— or —(OC₃H₆)_(y)—(OC₂H₄)_(x)—, and

x is in the range from 7 to 9 (preferably 8), and

y is in the range from 1 to 3 (preferably 1 to 2), and has a cloud pointat a temperature in the range from 64 to 70° C.,

The second surfactant (ii) is selected from a linear alkoxylated fattyalcohol and an oxo alcohol of the formula R₂-A-OH,

wherein R₂ is a linear or branched C₁₂ to C₁₅ alkyl or alkenyl group,

wherein A is —(OC₂H₄)_(x)—(OC₃H₆)_(y)— or —(OC₃H₆)_(y)—(OC₂H₄)_(x)—, and

x is in the range from 1 to 3 (preferably 2), and

y is in the range from 4 to 6 (preferably 5), and has a cloud point at atemperature in the range from 39 to 41° C.;

The surfactant system provides the detergent solution in useconcentration with low-foaming properties and makes it clear oressentially clear at a temperature in the range from 0 to 55° C.,preferably in the range from 10 to 50° C. and further preferred in therange from 16 to 40° C. Surfactant (i) has a cloud point at atemperature in the range from 64 to 70° C. and surfactant (ii) has acloud point at a temperature in the range from 39 to 41° C. The skilledperson will understand from the chemical structure of the respectivesurfactants that the turbidity (cloud point) of the surfactant (i) willbe measured in demineralized water while the cloud point of the othersurfactant (ii) will be measured in butyl diglycol (5 g of surfactant in25 ml of 25% aqueous butyl diglycol).

The fatty alcohols appropriate in the present invention can beexemplified by the alcohols obtained from the fatty acids as mentionedabove. Oxo alcohols generally represent a mixture of the linear alcoholand the alcohol which is branched with methyl in 2-position. Preferablythe alcohol has 10 to 16 and 12 to 15 carbon atoms, respectively.Technical mixtures may additionally contain proportions a differentnumber of carbon atoms.

It is particularly preferred that the sum of nonionic surfactant ispresent in a concentration ranging from 1 to 20 wt.-%, preferably from 1to 15 wt.-% and further preferred from 2 to 10 wt.-%

Enzymes

The enzyme system includes at least two different proteases and a lipaseand optionally an amylase. The purpose of the enzyme system is to breakdown adherent proteinaceous materials typically found on instrumentsafter use, into forms that are readily dispersed into a water-based washsolution. Proteins left in or on implements such as instruments thatcannot be mechanically scrubbed may prevent chemical disinfecting agentsto perform in an efficient way.

Any protease or mixture of proteases, from any source, can be employedin the enzyme system, provided that the selected enzyme system is stablein the desired pH range and compatible with the inventive composition.In a preferred embodiment two different proteases are of the serineendopeptidases type. While the enzyme may be obtained commercially in asolid or liquid form, the liquid form is preferred for greaterconvenience in dispersing the enzyme during preparation of theconcentrated cleaning solution of the invention and for complete waterdissolution of the enzyme.

Preferred protease enzymes are stable at least in a pH range of 5 to 8,and are obtained from bacterial strains and have sufficient activity pergram of enzyme protein to economically solubilize and remove proteinsfrom instruments during the desired cleaning cycle.

Suitable protease enzymes are, for example, the enzymes obtained fromBacillus subtilis, Bacillus licheniformis and Streptomyces griseus. Morepreferably, the enzyme is one or more of the commercially availableserine endoproteases. These enzymes preferably cleave protein links onthe carboxyl side of hydrophobic amino acid residues, but are capable ofcleaving most peptide links. They convert their substrates into smallfragments that are readily dissolved or dispersed into a wash solution.

Exemplary proteases are commercially available under the trade name ofEverlase® 16 L, Liquanase®, Savinase®, Esperase® (Novozymes) orPurafect® Prime L, Purafect L, Purafect Ox, Prosperase (Genencor) orBlap®.

Lipases are commercially available under the trade name Lipex®, orLipolase® (Novozymes).

Amylases may also be included in the cleaning and detergent compositionof the present invention for removing carbohydrate-based foreignmaterials from medical instruments to be cleaned. Amylase enzymes can beobtained from any suitable source, such as bacterial strains, barleymalt, certain animal glandular tissues and any others known to the art.Preferred amylase enzymes are stable in a pH range of 5 to 8, and areobtained from bacterial strains. Preferred types of amylases includethose which are referred to as alpha-amylases, beta-amylases,iso-amylases, pullulanases, maltogenic amylases, amyloglucosidases, andglucoamylases, as well as other amylases enzymes not particularlyelucidated here. These include also endo- and exo-active amylases. Suchamylases are commercially available under the trade name Purastar® ST,Purastar® HP AmL (Genencor), Stainzyme®, Duramyl®, Termamyl®, Termamyl®Ultra (Novozymes).

In a preferred embodiment the composition comprises 0.05 wt.-% to 10.0wt.-% of at least two different proteases, 0.05 wt.-% to 3.0 wt.-% oflipase and optionally 0.05 wt.-% to 3.0 wt.-% of amylase.

Enzyme Stabilizers

Preferred enzyme stabilizers include boron compounds or a calcium salts.More preferred, the enzyme stabilizers are a boron compound selectedfrom the group consisting of boronic acid, boric acid, borate,polyborate and combinations thereof and wherein the boron compound ispresent in a concentration ranging from about 0.2% to about 10% byweight. When calcium salts are used the calcium ion concentration is inthe range from 0.01 wt.-% to 3.0 wt.-%.

Water and Solvents

The concentrate composition preferably includes water ranging in aconcentration of 50 wt.-% or less, preferably of 20 wt.-% or less,further preferred of 15 wt.-% or less. The low amount of watercontributes to the stabilization of the enzymes during storage.

In addition to water, the composition may also include a solvent such asan alkanol or a polyol The alkanol is preferably soluble or misciblewith water and lipids, and comprises a C₁ to C₁₀ alkyl group that isstraight or branched, substituted or non-substituted. Useful alkanolsinclude short chain alcohols, such as C1-C8 primary, secondary andtertiary alcohols, e.g., methanol, ethanol, n-propanol, iso-propanol,and butanol. Particularly preferred alkanols include the various isomersof C3 alcohols, particularly iso-propanol. C1-C8 diols may also be usedin the alkanol constituent. The alkanol is present in a suitableconcentration, generally ranging from about 2 to about 10 percent, byweight, relative to the weight of the concentrate. Preferably, thealkanol concentration ranges from about 3 to about 10% by weight, and/orfrom about 7% to about 10%, by weight, of the enzyme-based aqueouscleaning compositions, i.e., the concentrate according to the invention.

The polyol is preferably an alkylene glycol, such as ethylene glycol orpropylene glycol. The polyols represented by the general formula (I) areexemplified by ethylene glycol, 1,2-propylene glycol, 1,3-propyleneglycol, glycerine, 1,4-butylene glycol and mixtures thereof, wherein1,2-propylene glycol is most preferred. The inventive compositioncontains water and/or the compound of formula (I) in an amount of from10 to 70 wt.-%, preferably from 30 to 65 wt.-%, wherein the watercontent preferably is 50 wt.-% or less, more preferred 20 wt.-% or less,further preferred 15 wt.-% or less, and the content of the compound offormula (I) preferably ranges from 20 to 55 wt.-%, more preferred from20 to 50 wt.-%, based on the whole composition.

Additional Adjuvants

The composition according to the present invention preferably is furthercomprising a metal corrosion inhibitor which is present in aconcentration ranging from about 0.050 to about 1 wt.-%.

The enzymes present in the composition may lead to increased foaming.Therefore, the composition preferably is further comprising an anti-foamcomponent, which further preferred is a silicone-based anti-foamcomponent.

The composition can also include an alkanolamine preferably selectedfrom the group consisting of monoalkanolamine, dialkanolamine,trialkanolamine, alkylalkanolamine, trialkylamine, triethanolamine andcombinations thereof, in a concentration ranging from about 1% to about10 wt.-%. The alkanolamine serves as pH adjustment.

Method of Use

The object of the present invention was also solved by a method forpreparing a detergent solution in use concentration which is intended tobe used for cleaning or pre-cleaning of surgical, medical or dentalinstruments or endoscopes by diluting the concentrate according to thepresent invention as outlined above with water to a percentage rangingfrom about 85 to about 0.5 wt.-%, wherein the detergent solution in useconcentration has low-foaming properties and is clear or is essentiallyclear at a temperature in the range from 0 to 55° C., preferably in therange from 10 to 50° C. and further preferred in the range from 16 to40° C.

Further, the present invention provides a method of cleaning orpre-cleaning surgical, medical or dental instruments, or endoscopes bycontacting the items with an effective amount of the liquid detergentcomposition according to the present invention as described above, for asufficient time to remove substantially all undesirable foreign matter,and then removing the liquid cleaning and detergent composition fromsaid instruments.

The method is carried out either manually or in ultrasonic applicationor as automated application in washers, preferably at a temperature inthe range from 0 to 55° C., further preferred at a temperature in therange from 10 to 50° C. and even further preferred at a temperature inthe range from 16 to 40° C.

In a further preferred method the effective amount of the liquidcleaning and detergent composition is from 0.5 wt.-% to 85 wt.-%,preferably from 0.5 wt.-% to 15 wt.-% in water.

According to the present invention the use dilution of the detergentsolution has low-foaming properties and is clear or is essentially clearat a temperature in the range from 0 to 55° C., preferably at atemperature in the range from 10 to 50° C. and further preferred at atemperature in the range from 16 to 40° C.

The present invention also provides the use of the liquid detergentcomposition according to the present invention as described above forcleaning surgical, medical or dental instruments or endoscopes.

Applications where enzymatic cleaning is desirable include the cleaningor pre-soaking of items or objects with metal parts or components thatmust be cleaned to remove organic or biological materials.

In addition to the above-mentioned advantages, the inventive cleaningsolution is substantially non-corrosive to metals such as stainlesssteel, anodized aluminium, aluminium, copper and brass which are oftenat risk of damages when soaked in previously available cleaningproducts.

The cleaning solution provided by the invention can be used for anysuitable purpose. Preferably, it is employed as a soak cleaner formedical and surgical instruments, dental hand piece, and the like, forboth human and veterinary practice. When used as a soak cleaner, it isapplied before the instruments are disinfected. It can also be readilyused as a concentrate to be added to ultrasonic baths and to automaticwasher, for cleaning more elaborate medical equipment.

When employed in automatic washing systems, it is important that thedetergent or surfactant component have low foam characteristics to avoidmachine malfunction due to excessive foam formation. The inventivecleaning and detergent solution is readily employed at elevatedtemperatures, as high as about 55° C.

Foreign matter to be cleaned or removed includes, for example,biological substances, e.g., tissue and blood. Foreign matter alsoincludes other materials such as lubricant, diagnostic and therapeuticcompositions, materials for pathology testing, medical or veterinaryresearch, and the like, remaining on instruments after these areemployed for their intended use. Foreign matter also includes pathogensincluding bacteria, viruses, and prions.

The term “instrument” is intended to be defined broadly, to include anyitems, objects, implements or devices for which the gentle removal oforganic or biological substances, such as protein, fats, carbohydratesand similar material is desirable. Simply by way of example and withoutlimitation, instruments are implements employed in patient or clientcontact (human or veterinary) during the practice of surgery, medicine,dentistry, podiatry, pathology for e.g., therapeutic, diagnostic and/orresearch purposes. Examples include, surgical instruments, e.g.,scalpels, probes, clamps, etc., endoscopes, operating room or dentalhandpieces, ventilation tubes, and the like. Surgical instruments andequipment inevitably pick up amounts of bio-burden on them after beingemployed in operations on humans or animals. Surgical instrumentsinclude for example rigid and flexible scopes, laparoscopic instruments,trays and anything that gets soiled with body fluids which result inthem having varying amounts of bio-burden on them after being so used.

Other devices that may be subjected to the inventive cleaning anddetergent composition also include miscellaneous other instrumentsand/or implements employed in cosmetic and beauty applications. Theseapplications include hair cutting, nail care, body art, skin piercing,collection of body fluids, e.g., blood, and blood separation andfractionation equipment, and similar such applications, that are toonumerous to mention. The inventive cleaning solutions are alsocontemplated to be useful for cleaning implements and items employed inthe food processing and pharmaceutical industries.

EXAMPLES Example 1 General Formulation of the Composition of the PresentInvention

The tests in respect to foam performance and cleaning efficacy wascarried out using liquid detergent compositions based on the generalcomposition as shown in table 1 (without enzymes), to which differentcombinations of enzymes have been added as shown in table 2. Theexamples shown in table 2 are based on the formulation shown in table 1.The percentage given for the enzymes in table refer to g of the actualproduct per 100 g of liquid base formulation. The final enzymeconcentration is indeed lower.

There are several products on the market based on enzymes and surfactantwhich are displaying different performances with regard to foamformation, turbidity and cleaning performance. A selection of theproducts on the market has been used as comparative examples forcomparison with the composition of the present invention, such ascompetitor product A (comprising protease, lipase, amylase and anionicsurfactant), competitor product B (comprising enzymes and nonionicsurfactant), competitor product C (comprising protease, lipase, amylaseand anionic surfactant), competitor product D (comprising protease,amylase, lipase, carbohydrase and nonionic surfactant), competitorproduct E (comprising protease, lipase, amylase and tensides),competitor product F (protease and nonionic surfactant).

TABLE 1 General formulation of the concentrate composition according tothe present invention without enzymes. component weight-% water 15.0glycol 47.5 corrosion inhibitor 0.5 preservative 0.5 alkanol 5.0 enzymestabilizers 2.2 alkanolamine 2.0 emulsifier 10.0 glycerol 4.5 defoamer(silicone based) 0.05 surfactant i) 4.5 C10-16 oxyalcohol 8 EO 1-2 POadduct surfactant ii) 3.5 C12-15 oxyalcohol 5 PO 2 EO adduct enzymes ad100.0

TABLE 2 The examples shown in this table are based on the formulation asdescribed in table 1. Lipase Protease [wt.-%] [wt.-%] Amylase [wt.-%]/test solution A B C D E A A B C D E 23 — — — — — — — — — — — 24 3 — — —— — — — — — — 25 — 3 — — — — — — — — — 26 — — 3 — — — — — — — — 27 — — —3 — — — — — — — 28 — — — — 3 — — — — — — 29 — 3 3 — — — — — — — — 30 — 3— — — 3 1.5 — — — — 31 — — 3 — — 3 1 — — — — 32 — 3 3 — — 3 1 — — — — 33— 3 — — — 3 2 — — — — 34 — 3 3 — — 3 2 — — — — 35 — 3 3 — — 3 — — — — —36 — 3 3 — — 2 — — — — — 37 — 3 3 — — 2 — 2 — — — 38 — 3 3 — — — — 2 — —— 39 — 2 2 — — 2 — — — — — 40 — 1 1 — — 1 — — — — — 41 — 2 2 — — 0.5 — —— — — 42 — 2 2 — — 1.5 — — — — — 43 — 2 2 — — 3 — — — — — 44 — 2 2 — — —— — — — — 45 — 2 2 — — 0.5 1 — — — — 46 — 2 2 — — 0.5 — 1 — — — 47 — 2 2— — 0.5 — — 1 — — 48 — 2 2 — — 0.5 — — — 1 — 49 — 2 2 — — 0.5 — — — — 150 — 2 2 — — 0.5 — — — — 0.5 64 — 2 2 — — 0.5 — — — — 0.25

Example 2 Testing the Blood Cleaning Efficacy

The products were tested regarding the efficiency of blood cleaning.Before the test the contact time is optimized with blood coupons(preparation see description below). Contact time in our test was set to10 minutes. The procedure is carried out as follows:

Stainless steel coupons are cleaned and degreased for preparation andthen weighed. 0.2 g of reactivated sheep blood is given onto a couponand spread on the surface so that a 3 mm frame stays clean. The couponsare then drying in a drying cabinet for 1 hour at 37° C. After coolingdown to room temperature the coupons are weighed again.

In order to perform the test 400 ml use solution of the detergentcomposition according to the present invention or of the respectivecompetitor product were added into a beaker. The test coupons wereimmersed in the detergent composition in a use concentration of 0.6% andleft for 10 min without agitation. After the contact time the couponswere removed and immersed in clear water for rinsing. Then the testcoupons were dried at ambient temperature over night.

The test was evaluated visually and gravimetrically. The weight of thecoupons with dried blood minus the weight of clean coupons is equal tothe blood load; the weight of cleaned coupons minus the weight of theclean coupons is equal to the blood load after cleaning, namely theremaining blood load. Using this approach the cleaning efficacy can becalculated. The result in form of the cleaning efficacy is given inpercent. The tests were repeated minimum three times.

The experiments were carried out at a temperature in the range of 20 to23° C. and show that the blood cleaning efficacy increases when proteaseis added (test solutions No: 24-28) compared to the test solution havingthe base formulation without any enzymes (No: 23). The addition of asecond (different) protease (test solution No: 29, comprising twodifferent proteases) results in a further increased blood cleaningefficacy. The test solutions having one or two different proteases andin addition lipase and/or lipase and amylase (test solutions No: 30-43and 45-64) also have increased blood cleaning efficacy compared to testsolutions only comprising one protease as enzyme.

Example 3 Testing the Blood Cleaning Efficacy at Different Temperatures

The products were tested regarding the efficiency of blood cleaning atdifferent temperatures, namely at 20-23° C., 40° C. and 55° C. The testmethod is the same as in example 2. The results are shown in table 3.The test solution 23 represents the base formulation without enzymes asshown in table 1. The test solutions A, B, C, D and F representcompetitor products as described above. The solution depicted with thenumber “64” represents the formulation according to the presentinvention comprising enzymes as listed in table 2.

TABLE 3 Assaying different test solutions and comparative examples inrespect to the Blood Cleaning Efficacy at temperatures of 20-23° C., 40°C. and 55° C. for 10 min, n = 3. test solution/ relative averagecleaning + = invention (usage efficacy in %, n = 3 − = comparativeconcentration [%]) 20° C. 40° C. 55° C. example 23/(0.6) 98.7 93.3 93.8− 64/(0.6) 99.6 95.1 97.9 + C/(0.5) 100.0 95.7 95.5 − A/(0.8) 98.6 95.894.5 − B/(1.0) 99.2 95.7 95.4 − F/(1.0) 97.9 95.4 93.6 − D/(0.4) 99.295.3 93.5 −

The experiments show that the composition according to the presentinvention No: 64 has comparable well blood cleaning efficacy and atleast at a temperature of 55° C. has improved blood cleaning efficacycompared to other compositions. Therefore, the compositions according tothe present invention have advantageous properties over a wider rangecompared to the compositions of the state of art. 1

Example 4 Testing the Foaming Performance

Description of the test in respect to the foaming performance (shakingmethod)

The use solution (100 ml) is filled into a 250 ml measuring cylinder.The cylinder is closed with a PE plug and tilted 20 times (to upsidedown). After time points 0 min, 1 minute and after 5 minutes the foamheight in the measuring cylinder is measured with a ruler and documentedin “mm”. This test is repeated minimum three times.

Before the test the measuring cylinder has to be cleaned and rinsed withethanol for degreasing; and after that rinse with demineralized water.

Description of the Test in Respect to the Foaming Performance inIntensive Foaming Machine

The foaming performance of the products in use solution is measured withmechanical action in dependency of the temperature. The intensive foammachine holds a volume of 40 L. The product is added according to thedesired use concentration. After the product has been added themechanical action is started. The mechanical action simulates thesituation in an automated washer. The starting temperature is between20° and 22° C. After the mechanical action has been running for 1 minutethe machine is stopped and the foam height documented. Then theagitation is started again and the heating is switched on. In intervalsof 2° C. the machine is stopped and the foam height documented. Thetemperature is raised up to 55° C. to simulate a washer. The foam heightis documented in mm. The results are summarized in table 4.

TABLE 4 Foam performance of test solution and comparative examples. foamperformance (cm) test solution 0 min 1 min 5 min competitor product A 76 5 (0.8%) competitor product B 7.5 3 0.5 (1.0%) competitor product C 31 1 (0.5%) competitor product D 7 5 4 (0.425%) competitor product E 1513 9 (1.0%) competitor product F 6.5 2 1 (1.0%) 64 (invention) 6 2.5 1.5

The experiments show that the composition according to the presentinvention No: 64 has low-foaming properties compared to most of othercompositions. Therefore the composition of the present invention isparticularly useful for manual cleaning of surgical instruments andendoscopes.

In respect to testing the foaming performance in intensive foamingmachine method the results show ideal properties of the compositionaccording to the present invention. The foam height was at a temperaturefrom about 20° C. to 42° C. was between 300 to 400 mm, while at atemperature of 44° C., 46° C., 48° C., 50° C. and 52° C. the foam heightwent down and was 250, 210, 190, 120 and 100 mm, respectively, while ata temperature of 54° C. the foam height was less 20 mm or less.

Example 4 Description of the Assay of the Cloud Point of Products andUse Concentrations

The method is used for the determination of the cloud point in liquidproducts. The temperature describes the temperature range in which thenonionic surfactant in diluted concentration becomes harder soluble andby that low foaming.

The cloud point is understood as a temperature where the liquid productgets cloudy. The procedure is carried out as follows:

The use concentration is filled into a 150 ml beaker. The temperature israised slowly and monitored with a thermometer. The temperature at whichthe product solution shows the first cloudiness during heating isdocumented. The solution as well as the temperature are observed duringcooling and the temperature is documented where the solution firstbecomes clear again. The cloud point is determined with an accuracy of0.5° C. The results are summarized in table 5.

TABLE 5 The table summarizes the results of the measurements of thecloud points/turbidity of diverse competitor products and thecomposition according to the present invention. test solution cloudpoint in demineralized water competitor product A (0.8%) n.d. competitorproduct B 35° C. (1.0%) competitor product C n.d. (0.5%) competitorproduct D (0.43%) 40.5° C.  competitor product E n.d. (1.0%) competitorproduct F 26° C. (1.0%) invention >55° C. 

The results show that the liquid detergent composition of the presentinvention gives a cleaning solution which is not cloudy up to atemperature of 55° C. Due to these characteristics the composition ofthe present invention is particularly useful for manual and ultrasonicapplication when cleaning surgical instruments and endoscopes.

1. A liquid detergent concentrate composition comprising: a) 1 to 30wt.-% of a surfactant system comprising at least two different nonionicsurfactants, wherein the first surfactant has a cloud point at atemperature in the range from 64 to 70° C., and the second surfactanthas a cloud point at a temperature in the range from 39 to 41° C.; b) atleast two different proteases; c) 10 to 70 wt.-% water; and d) an enzymestabilizer wherein when the concentrate is diluted to form a usecomposition the use composition is low-foaming and essentially clear ata temperature in the range from 0 to 50° C.
 2. The composition accordingto claim 1, wherein the surfactant (i) is selected from the groupconsisting of a linear alkoxylated fatty alcohol and an oxo alcohol ofthe formula R₁-A-OH, wherein R₁ is a linear or branched C₁₀ to C₁₆ alkylor alkenyl group, wherein A is —(OC₂H₄)_(x)—(OC₃H₆)_(y)— or—(OC₃H₆)_(y)—(OC₂H₄)_(x)—, and x is from 7 to 9, and y is from 1 to 3,and the surfactant (ii) is selected from the group consisting of alinear alkoxylated fatty alcohol and an oxo alcohol of the formulaR₂-A-OH, wherein R₂ is a linear or branched C₁₂ to C₁₅ alkyl or alkenylgroup, wherein A is —(OC₂H₄)_(x)—(OC₃H₆)_(y)— or—(OC₃H₆)_(y)—(OC₂H₄)_(x)—, and x is from 1 to 3, and y is from 4-6. 3.The composition of claim 1, wherein the sum of nonionic surfactant ispresent in a concentration ranging from 1 to 20 wt.-%.
 4. Thecomposition of claim 1, wherein each of the different at least twoproteases is a serine endopeptidase.
 5. The composition of claim 1,wherein the water is present in a concentration of 50 wt.-% or less. 6.The composition of claim 1, wherein the enzyme stabilizer is selectedfrom the group consisting of a boron compound and a calcium salt.
 7. Thecomposition of claim 1, wherein the enzyme stabilizer is a boroncompound selected from the group consisting of boronic acid, boric acid,borate, polyborate and combinations thereof and wherein the boroncompound is present in a concentration ranging from about 0.2% to about10% by weight.
 8. The composition of claim 1, further comprising analkanolamine selected from the group consisting of monoalkanolamine,dialkanolamine, trialkanolamine, alkylalkanolamine, trialkylamine,triethanolamine and combinations thereof, in a concentration rangingfrom about 1% to about 10 wt.-%.
 9. The composition of claim 1, whereinthe composition further comprises at least one alkanol, wherein thealkanol is soluble or miscible with water and lipids, and comprises a C₁to C₁₀ alkyl group that is straight or branched, substituted ornon-substituted.
 10. The composition of claim 1, further comprising ametal corrosion inhibitor present in a concentration ranging from about0.050 to about 1 wt.-%.
 11. The composition of claim 1, furthercomprising a silicone-based anti-foam component.
 12. The composition ofclaim 1, having a pH in the range from 5.5 to 9.0.
 13. The compositionof claim 1, further comprising an additional enzyme selected from thegroup consisting of lipase, amylase and mixtures thereof.
 14. A methodof cleaning instruments or endoscopes comprising: (a) contacting theinstrument or endoscope with an effective amount of a liquid detergentconcentrate composition in water, the detergent composition comprising:1 to 30 wt.-% of a low-foam surfactant system comprising at least twodifferent nonionic surfactants, wherein the first surfactant has a cloudpoint at a temperature in the range from 64 to 70° C., and the secondsurfactant has a cloud point at a temperature in the range from 39 to41° C.; at least two different proteases; 10 to 70 wt.-% water; andenzyme stabilizers, wherein when the concentrate is diluted to form ause composition the use composition is low-foaming and essentially clearat a temperature in the range from 0 to 55° C.; and (b) removingsubstantially all undesirable foreign matter from the instrument orendoscope; and (c) rinsing the instruments or endoscope.